Acetylcholine receptor is a protein that spans the plasma membrane and forms a gated channel for cations between the cytoplasm and the exterior of the muscle cell. The portion of this channel that passes through the membrane itself is formed by a ring of five, juxtaposed, hydrophobic domains, one from each of the five subunits. It is the long-term objective of this project to define the precise molecular structure of each of these domains and, thereby, that of the ring itself and the channel for the cations. The four membrane-spanning sequences in each of the four subunits of acetylcholine receptor have been identified, and Kendrew-Watson wire models have been constructed of the four membrane-spanning helices of the Gamma subunit. These four hilices have been aligned to create a hypothetical molecular model for the membrane-spanning domain of this protein. This model contains two intrasubunit disulfide bonds within the bilayer. These two disulfides will be sought within the native structure of acetylcholine receptor through chemical experiments performed on the purified protein. Peptides containing these disulfides will be isolated and identified by methods that have been used successfully to purify the membrane-spanning portions of (Na+ + K+)-ATPase. If these disulfides both exist, there is a high probability that the hypothetical model is correct. If they do not, other intrasubunit disulfides may occur within the membrane-spanning domains of other subunits in the native structure, and these will also be sought. If every one of the eleven unique cysteine residues within the membrane, however, are free, cross-linking agents will be attached to any of them that are accessible from the alkane core of the bilayer. Covalent cross-links between these cysteines and neighboring amino acid side chains will be formed, and the products of these cross-linking reactions will be purified and identified. If one or two of any of the foregoing chemical experiments provide positive results, they should define a unique molecular structure for the membrane-spanning domain of at least one of the four subunits of acetylcholine receptor. This, however, will define the structure of each of the four domains from each of the four subunits because every subunit is necessarily folded in the same, superimposable pattern.